Project Summary/Abstract It has long been recognized that chronic exposure to stressful adverse life experiences confers susceptibility to developing post-traumatic stress disorder (PTSD), a stressor and trauma-related disorder characterized by intense fearful memory formation. Many of the long-lasting symptoms associated with PTSD can be attributed to an impaired ability to discriminate between cues associated with threat from those associated with safety. Chronic stress is believed to play a key role in the etiology of PTSD by altering the brain's fear memory system in a way that increases generalization of fear to `safe' cues that are unrelated to the trauma. Recent findings have suggested that memories of fear and safety are encoded and expressed via an elaborate pattern of communication between the medial prefrontal cortex (mPFC) and the basolateral amygdala (BLA), with BLA activity being suppressed by input from the mPFC during periods of recognized safety. Moreover, evidence suggests that basal forebrain (BF) cholinergic inputs to the mPFC play a critical role in facilitating the detection and encoding of certain types of learned cues. It remains unknown, however, whether cholinergic input to the mPFC is involved in the encoding of safety cues, and no studies have conducted a circuit-level analysis to examine how a history of chronic stress regulates dynamic network activity between the BF, mPFC and BLA. In the present proposal, we will first use behavioral and immunocytochemical techniques to characterize the effects of chronic stress on fear and safety discrimination and activity within the basal forebrain cholinergic system in male and female mice (Aim I). Next, we will use multisite neurophysiological recordings in awake-behaving male and female mice to systematically examine how a history of chronic stress regulates dynamic patterns of neural activity between the BF, mPFC and BLA during fear and safety discrimination (Aim II). Finally, we will use optogenetic methods to manipulate activity in the BF-mPFC pathway to determine how cholinergic input contributes to fear discrimination learning and how it might be impacted by a history of chronic stress in both sexes (Aim III). Our findings have the potential to significantly broaden our current understanding of how exposure to chronic stress modulates fear discrimination and the adaptive use of safety cues at the neural circuit level, findings which may have relevance for the treatment of disorders such as PTSD.